This project delineates biochemical and pharmacological properties of sigma-1 receptors. Sigma-1 receptors are one-transmembrane proteins at the endoplasmic reticulum (ER) that bind neurosteroids, dextrobenzomorphans, and certain psychostimulants such as cocaine. In this study, we found that sigma-1 receptors specifically target lipid storage sites (lipid droplets) on the endoplasmic reticulum by forming a distinct class of lipid microdomains. Both endogenously expressing sigma-1 receptors and transfected C-terminally EYFP-tagged sigma-1 receptors (Sig-1R-EYFP) target unique ?ring-like? structures associated with endoplasmic reticulum reticular networks in NG108-15 cells. The ?ring-like? structures contain neutral lipids and are enlarged by the oleate treatment, indicating that they are endoplasmic reticulum-associated lipid droplets (ER-LD). Sigma-1 receptors colocalize with caveolin-2, a cholesterol-binding protein in lipid rafts on the ER-LD, but not with ADRP, a cytosolic lipid droplet (c-LD) specific protein. When the double-arginine ER retention signal on the N-terminus of sigma-1 receptors is truncated, sigma-1 receptors no longer exist on ER-LD, but predominantly target c-LD which contain ADRP. Sigma-1 receptors on ER-LD form detergent-resistant raft-like lipid microdomains, the buoyancy of which is different from those of plasma membrane lipid rafts. (+)Pentazocine causes sigma-1 receptors to disappear from the microdomains. N-terminally EYFP-tagged Sigma-1 receptors (EYFP-Sig-1R) failed to target ER-LD. EYFP-Sig-1R-transfected cells showed an unrestricted distribution of neutral lipids all over the endoplasmic reticulum network, decreases in c-LD and cholesterol in plasma membranes, and the bulbous aggregation of endoplasmic reticulum. Thus, sigma-1 receptors are unique endoplasmic reticulum proteins that regulate the compartmentalization of lipids on the endoplasmic reticulum and their export from the endoplasmic reticulum to plasma membrane and c-LD.